Analysis of the protective capacity of SAG1 and SAG2 subunit vaccines in BALB/c mice

by Yang, Chung-Dar

Abstract (Summary)

IV
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SAG1 and SAG2 are important surface molecules of T. gondii for the
invasion of tachyzoites into host-cells. Previous studies have been
demonstrated they are good candidates for development of vaccines
against toxoplasmosis. Therefore, we used SAG1 and SAG2 to generate
subunit vaccines and evaluated the protective capacity in BALB/c mice.
Anti-idiotype IgG (aId-IgG) with the SAG2 internal image was prepared
from anti-SAG2 monoclonal antibody in accordance with the network
theory. Lymphocytes from mice immunized subcutaneously twice with 2,
4 or 8 µg aId-IgG showed great proliferations and secreted high level of
IFN-£^, which is an important cytokines secreted by Th1 cells. After
challenged subcutaneously with 1¡Ñ103 tachyzoites, the highest survival
rate reached 88%. Further, SAG1 and SAG2 genes were respectively
cloned and recombinant proteins rSAG1 and rSAG2 were prepared.
Lymphocytes from mice immunized intraperitoneally twice with rSAG1
or rSAG2 displayed apparently Th1-associated responses, while
lymphocytes from mice immunized subcutaneously twice with rSAG1 or
rSAG2 did not. Mice immunized intraperitoneally twice with rSAG1 or
rSAG2 had a survival rate of 64% which was higher than those mice
immunized subcutaneously with rSAG1 or rSAG2. When mice
immunized intraperitoneally twice with rSAG1 mixed with rSAG2,
survival rate was even higher (71%). Therefore, mixed antigens induced
higher protection. Subsequently, SAG1 gene was fused with SAG2 gene
and a recombinant protein rSAG1/2 was expressed from the fused gene.
Th1-associated responses were detected from lymphocytes of mice
immunized intraperitoneally twice with 10 µg rSAG1/2. Interestingly,
80% rSAG1/2-immunized mice survived and it was extremely higher
V
than rSAG1- or rSAG2-immunized mice (64%). In an attempt to
stimulate immune responses against T. gondii infections in the mucosal
system, we chose heat-labile enterotoxin (LT) secreted from toxigenic E.
coli as the stimulator. LTA2B and LTB genes were respectively cloned and
then rLTA2B and rLTB were obtained. Moreover, LTA2B gene or LTB
gene fused with SAG1 and SAG2 genes was performed and then
recombinant proteins rLTA2B-SAG2/1 and rLTB-SAG1/2 were prepared.
Subsequently, mice were immunized intranasally twice with
rLTA2B-SAG2/1, rLTB-SAG1/2, rLTA2B mixed with rSAG1/2, or rLTB
mixed with rSAG1/2. A strong protection (67%) was shown by the group
of mice immunized intranasally with 10 µg rLTB-SAG1/2 or 4 µg rLTB
mixed with 10 µg rSAG1/2. rLTB, rather than rLTA2B, will be a better
candidate for stimulation the mucosal system. In summary, different
survival rates caused by antigens prepared in the study may be attributed
to many factors such as the treatment, the preparation and the dose of
antigen. The highest survival rate is caused by aId-IgG (88%); the second
is shown by rSAG1/2 (80%); the third is resulted from rSAG1mixed
rSAG2 (71%). Although the survival rate raised by rLTB-SAG1/2 or
rLTB mixed rSAG1/2 is slightly low (67%), these data demonstrate
stimulators such as LT could induce anti-Toxoplasma immune response
of the mucosal system. Antigens capable of inducing higher survival rates
in mice should be worthy of further investigation for searching better
vaccine candidates.